Protease activated receptor 2 controls myelin development, resiliency and repair

Abstract

Oligodendrocytes are essential regulators of axonal energy homeostasis and electrical conduction and emerging target cells for restoration of neurological function. Here we investigate the role of protease activated receptor 2 (PAR2), a unique protease activated G protein-coupled receptor, in myelin development and repair using the spinal cord as a model. Results demonstrate that genetic deletion of PAR2 accelerates myelin production, including higher proteolipid protein (PLP) levels in the spinal cord at birth and higher levels of myelin basic protein and thickened myelin sheaths in adulthood. Enhancements in spinal cord myelin with PAR2 loss-of-function were accompanied by increased numbers of Olig2- and CC1-positive oligodendrocytes, as well as in levels of cyclic adenosine monophosphate (cAMP), and extracellular signal related kinase 1/2 (ERK1/2) signaling. Parallel pro-myelinating effects were observed after blocking PAR2 expression in purified oligodendrocyte cultures, whereas inhibiting adenylate cyclase reversed these effects. Conversely, PAR2 activation reduced PLP expression and this effect was prevented by brain derived neurotrophic factor (BDNF), a pro-myelinating growth factor that signals through cAMP. PAR2 knockout mice also showed improved myelin resiliency after traumatic spinal cord injury and an accelerated pattern of myelin regeneration after focal demyelination. These findings suggest that PAR2 is an important controller of myelin production and regeneration, both in the developing and adult spinal cord.

A Western diet impairs CNS energy homeostasis and recovery after spinal cord injury: Link to astrocyte metabolism

“…All reagents were purchased from Gibco (Invitrogen, Carlsbad, CA, USA). After 10–12 d in vitro (DIV), mixed glial cultures were shaken overnight (225 RPM) to remove microglia and oligodendrocytes as previously detailed ( Burda et al, 2013 ; Yoon et al, 2017 ). Astrocytes were tryp-sinized (0.025%) and plated at a density of 450,000 cells/well on poly-L-lysine- (Sigma-Aldrich) coated 6-well plates and grown for 2–3 d in serum-free Neurobasal A media containing 1% N2, 2% B27, 100 U/ml P/S, 1 mM sodium pyruvate, 0.45% glucose, 0.5% Glutamax, 0.1% BSA, and 50 μM β-mercaptoethanol.…”

Blocking the Thrombin Receptor Promotes Repair of Demyelinated Lesions in the Adult Brain

“…Images throughout the site of focal demyelination were taken under oil immersion at 60ϫ (Olympus BX51 microscope) and montaged to visualize the complete lesion in Adobe Photoshop. The total lesion area was measured and counts of remyelinated axons (defined as lightly stained and thin myelin sheaths) were expressed as the number of remyelinated axons per square millimeter of lesion (Yoon et al, 2017).…”

“…Immunochemical analyses of cellular changes across the lysolecithin and cuprizone models was performed on paraffin embedded materials as previously detailed (Radulovic et al, 2016;Yoon et al, 2017). Briefly, sections were deparaffinized followed by antigen retrieval (sodium citrate,10 mM, pH 6.0), blocked with serum with or without 0.25% Triton X-100 followed by incubation with primary antibodies overnight.…”

“…Primary oligodendrocyte or astrocyte cultures were purified from mixed glial cultures prepared from the cortices of postnatal day 1 to 3 mice as described previously (Radulovic et al, 2016;Yoon et al, 2017). In all cases, mixed glia were grown in Dulbecco's modified Eagle's medium (DMEM) (11960-044, Invitrogen), containing 1 mM sodium pyruvate (11360070, Corning), 20 mM HEPES (15630-080, Invitrogen), 10% heatinactivated fetal bovine serum (Atlanta Biologicals), 100 U/ml penicillin and 100 g/ml streptomycin (15140122, Life Technologies).…”

Microenvironment Imbalance of Spinal Cord Injury

“…Although spared and regenerated axons are myelinated, the conductive function of the axons does not change 49 . Thus, many studies demonstrated SCI repair and recovered spinal cord function through the restoration of the myelin sheath 50 – 52 . Our team transplanted autologous activated Schwann cells into the spinal cord, and we found relatively complete restoration of the myelin sheath and improved microenvironment balance 53 , and another study demonstrated similar results when co-transplanting human umbilical cord mesenchymal stem cells and human Schwann cells 54 .…”